Expression of CD56, a marker of natural killer (NK) cells, in acute lymphocytic leukemia (ALL) is rare and, to date, has been described only in non-B lineage ALL. Among 194 patients with CD56 analysis on the ongoing Eastern Cooperative Oncology Group (ECOG) ALL trial, E2993, 6 cases of CD56+ ALL were found (3.1%) with a median of 95% of blast cells expressing CD56, compared with a median of 1% of blast cells in CD56- ALL (P = 0.0001). FAB-L2 characteristics dominated, without granulation. Blast cells from four CD56+ patients expressed T-cell antigens at variable levels of maturation. A clonal rearrangement of the T-cell receptor beta (TCRbeta) gene was detected only in one patient. TCRbeta variable gene usage studies in this and one other CD56+ ALL patient demonstrated a significantly perturbed usage pattern in both patients when compared with control lymphocytes. The two remaining cases typed as early pre-B ALL (CD19+, CD10+), with one case co-expressing CD7. Cytogenetically, 4 patients were normal, 1 complex abnormal, and 1 Philadelphia chromosome positive. Epstein-Barr virus (EBV) sequences were detected in one T- and both B-lymphoid cases. Our data suggest that CD56 is expressed at a precursor stage common to the T- and the B-cell lineage.